摘要
目的研究维生素D(VD)对大鼠肾上腺嗜铬细胞瘤细胞系PC12细胞中神经酰胺(Cer)水平的影响。方法采用C2-Cer(20μmol/L)刺激PC12细胞,MTT法检测Cer对PC12细胞增殖的影响;高效液相色谱串联质谱法(HPLC-MS)测定VD(50或0 nmol/L)处理的PC12细胞中C16:0-Cer的含量。qRT-PCR法测定VD(50或0 nmol/L)处理的PC12细胞中Cer代谢相关酶mRNA的表达;Western blotting法检测VD(200、100、50、25、12.5和0 nmol/L)处理后,PC12细胞中SPTLC1和Cers6蛋白的相对表达量。结果 C2-Cer能抑制PC12细胞的活性,IC50为2.800μmol/L。与0 nmol/L VD相比,50 nmol/L VD降低了PC12细胞中C16:0-Cer的含量[(19.37±0.61)ng/m L vs(16.50±0.10)ng/m L,P=0.010],升高了SPTLC1 mRNA的表达水平(P=0.001)和Cers6mRNA的表达水平(P<0.001),降低了SPTLC1蛋白的相对表达量(P=0.032),升高了Cers6蛋白的相对表达量(P=0.020)。结论 VD降低PC12中Cer的含量,可能与降低SPTLC1蛋白表达有关。
Objective To invastigate the effect of vitamin D（VD） on the content of ceramide（ Cer） in rat adrenal pheochromocytoma PC12 cells. Methods MTT was performed to verify the effect of short-chain C2-Cer（ 20 μmol/L） on cell proliferation. High performance liquid chromatography tandem mass spectrometry（ HPLC-MS） was performed to detect the concentration of C16： 0-Cer in PC12 cells stimulated with VD（ 50 or 0 nmol/L）. qRT-PCR was performed to measure the mRNA expression levels of enzymes which were related to the metabolism of Cer in PC12 cells after being stimulated with VD（ 50 or 0 nmol/L）. Western blotting was performed to measure the protein levels of SPTLC1 and Cers6 in PC12 cells that were stimulated with VD（ 200,100,50,25,12. 5 and 0 nmol/L）. Results Activity of PC12 cells was inhibited by C2-Cer with IC50 being 2. 800 μmol/L. Compared with VD at 0 nmol/L,the content of C16： 0-Cer in PC12 cells was decreased by VD at 50 nmol/L [（ 19. 37 ± 0. 61） ng/m L vs（ 16. 50 ± 0. 10） ng/m L,P= 0. 010],the expressions of SPTLC1 mRNA（ P = 0. 001） and Cers6 mRNA（ P〈0. 001） in PC12 cells were increased,the protein expression of SPTLC1 in PC12 cells was reduced（ P = 0. 032） and the protein expression of Cers6 was increased（ P = 0. 020）. Conclusion The concentration of C16： 0-Cer is decreased in PC12 cells that were treated with VD,and it may be related to the decrease of protein expression of SPTLC1.
出处
《山东大学学报（医学版）》
CAS
北大核心
2017年第12期13-17,23,共6页
Journal of Shandong University:Health Sciences
基金
国家自然科学基金(81271319)
